Unusually efficient CUG initiation of an overlapping reading frame in <i>POLG</i> mRNA yields novel protein POLGARF.

Abstract

While near-cognate codons are frequently used for translation initiation in eukaryotes, their efficiencies are usually low (<10% compared to an AUG in optimal context). Here, we describe a rare case of highly efficient near-cognate initiation. A CUG triplet located in the 5' leader of <i>POLG</i> messenger RNA (mRNA) initiates almost as efficiently (∼60 to 70%) as an AUG in optimal context. This CUG directs translation of a conserved 260-triplet-long overlapping open reading frame (ORF), which we call <i>POLGARF</i> (<i>POLG</i> Alternative Reading Frame). Translation of a short upstream ORF 5' of this CUG governs the ratio between POLG (the catalytic subunit of mitochondrial DNA polymerase) and POLGARF synthesized from a single <i>POLG</i> mRNA. Functional investigation of POLGARF suggests a role in extracellular signaling. While unprocessed POLGARF localizes to the nucleoli together with its interacting partner C1QBP, serum stimulation results in rapid cleavage and secretion of a POLGARF C-terminal fragment. Phylogenetic analysis shows that <i>POLGARF</i> evolved ∼160 million y ago due to a mammalian-wide interspersed repeat (MIR) transposition into the 5' leader sequence of the mammalian <i>POLG</i> gene, which became fixed in placental mammals. This discovery of <i>POLGARF</i> unveils a previously undescribed mechanism of de novo protein-coding gene evolution.